Transposable elements mediate rapid genetic change by creating allelic diversity and "hot spots" for recombination and translocations. In prokaryotes, including organisms causing human disease, transposons move antibiotic resistance genes between species. In eukaryotes, transposons generate allelic diversity on an evolutionary time scale or within a single organism. The goal of the proposed work is to understand how Mutator transposable element activities are regulated with the life cycle of its natural host, maize. Mutator (Mu) element transposons mobilized by MuDR produce the highest forward mutation frequency of any eukaryotic transposon. Mutator exhibits two forms of developmental regulation: [1] late timing as elements are mobile primarily after cell fate determination, and [2] two transposition mechanisms with excision in the soma and replicative insertion in the germinal cells and gametes. How do the diverse transcripts and potential proteins encoded by MuDR mediate developmental timing and the two transposition mechanisms? Carefully staged tissues will be analyzed for product appearance and abundance in several Mutator lines with different regulatory behaviors and in transgenic maize expressing myc-tagged MUR proteins. The possible contribution of antisense transcripts to developmental regulation will be analyzed by expressing sense and antisense transcripts to developmental regulation will be analyzed by expressing sense and antisense RNA from heat shock promoters in transgenic maize. Mutator activities will be reconstructed in transgenic plants expressing individual MuDR-encoded products; marked Mu elements (containing a pBluescript plasmid) will allow recovery of new insertions to distinguish between "cut only" "cut and paste" and "cut and paste with gap repair" models of somatic excision; the marked elements will be used to define the timing and frequency of Mu element amplification during replicative transposition in the germinal cells and gametes. In vitro biochemical studies will examine the DNA binding properties and enzymatic activities of the transposes encoded by MuDR to determine if different transposases are required for "cut and paste" and replicative transposition.